Enzyme immunoassays have been a very successful type of homogeneous immunoassay. A number of such assays have been based on the ability of fragments of .beta.-galactosidase to complement each other and form active enzyme. In particular, a .beta.-galactosidase enzyme donor (ED) combines with a .beta.-galactosidase enzyme receptor (EA) to form active .beta.-galactosidase enzyme. Conjugating a small analyte or an analyte analogue to the ED at certain sites does not affect the complementation of ED to EA or the rate of .beta.-galactosidase catalyzed activity. However, when the ED-analyte conjugate is bound by anti-analyte antibody, complementation and the enzyme-catalyzed reaction rate during the initial phase of the reaction is reduced. This reduction in enzyme-catalyzed reaction rate has been used to quantify analytes in a situation where both the ED-analyte conjugate present in an assay medium and the analyte present in the sample compete for anti-analyte antibody prior to the addition of EA. The .beta.-galactosidase catalyzed reaction rate increases as the amount of analyte present in the sample increases, because more analyte in the sample reduces interaction of the ED-analyte conjugate and anti-analyte antibody, allowing more of the ED-analyte conjugate to react with the EA to form active .beta.-galactosidase enzyme.
This technology has generally been limited to low molecular weight analytes. The present invention provides a method capable of measuring high molecular weight analytes.